Polymerase chain reaction amplification of a portion of the genome of both rapidly growing mycobacteria and nocardia, followed by restriction fragment length polymorphism (RFLP) analysis of the amplification products, is being evaluated to assess use of these procedures in the diagnostic laboratory. The technique has already proven useful in preliminary identification of these organisms within a few days of their isolation (compared with the month or more required for conventional identification with biochemical testing). In addition, these molecular procedures allow more accurate discrimination among species and subspecies than is possible with biochemical testing. We are correlating results obtained with phenotypic and molecular methods. For the nocardia, we will investigate the possible use of other primer pairs to ensure that amplification of the portion of the genome with which we are working can indeed be obtained from all species in the genus Nocardia. Results of the work performed to date have included the demonstration of (1) the unreliability of the conventional salt tolerance test for distinguishing among some of the species of rapidly growing mycobacteria and (2) the existence of two distinct RFLP patterns within one of the more commonly isolated species of Nocardia.